Fractalkine and CX3CR1 are involved in the recruitment of intraepithelial lymphocytes of intrahepatic bile ducts

Fractalkine is a chemokine with both chemoattractant and cell-adhesive functions, and in the intestine it is involved with its receptor CX3CR1 in the chemoattraction and recruitment of intraepithelial lymphocytes. We examined the pathophysiological roles of fractalkine and CX3CR1 in normal and diseased bile ducts. Expression of fractalkine and CX3CR1 were examined in liver tissues from patients with primary biliary cirrhosis (17 cases) and controls (9 cases of primary sclerosing cholangitis, 10 cases of extrahepatic biliary obstruction, 20 cases of chronic viral hepatitis C, and 18 cases of histologically normal livers). Expression of fractalkine in biliary epithelial cells (BECs) in response to cytokine treatments was examined using a human cholangiocarcinoma cell line (HuCC-T1) and human intrahepatic BEC line. The chemotaxis of CX3CR1-expressing monocytes (THP-1) toward fractalkine was assayed using chemotaxis chambers. Fractalkine messenger RNA/protein were expressed on BECs of normal and diseased bile ducts, and their expression was upregulated in injured bile ducts of primary biliary cirrhosis. CX3CR1 was expressed on infiltrating mononuclear cells in portal tracts and on CD3(+), CD4(+), and CD8(+) intraepithelial lymphocytes of injured bile ducts in primary biliary cirrhosis. Fractalkine messenger RNA expression was upregulated in two cultured BECs on treatment with lipopolysaccharide and Th1-cytokines (interleukin 1beta, interferon gamma, and tumor necrosis factor alpha). THP-1 cells showed chemotaxis toward fractalkine secreted by cultured cells. In conclusion, Th1-cytokine predominance and lipopolysaccharide in the microenvironment of injured bile ducts resulting from primary biliary cirrhosis induce the upregulation of fractalkine expression in BECs, followed by the chemoattraction of CX3CR1-expressing mononuclear cells, including CD4(+) and CD8(+) T cells, and their adhesion to BECs and the accumulation of biliary intraepithelial lymphocytes.     

Innervation of intrahepatic bile ducts and peribiliary glands in normal human livers, extrahepatic biliary obstruction and hepatolithiasis. An immunohistochemical study

Innervation of the intrahepatic biliary tree was examined in human normal livers, extrahepatic biliary obstruction and hepatolithiasis. Nerve fibers were immunohistochemically identified on formalin-fixed and paraffin-embedded sections by antibodies to S-100 protein (S-100) and neuron specific enolase (NSE). S-100 and NSE-immunoreactive nerve fibers were present in the walls of intrahepatic large, medium-sized and septal bile ducts as well as in peribiliary glands. Some nerve fibers were in close contact with epithelia of bile ducts and peribiliary glands. Serial section observations showed that the nerve fibers arising from nerve bundles approached, and came to lie in close contact with epithelia of the bile ducts and peribiliary glands. Nerve fibers were sparse around the interlobular bile ducts and bile ductules. These immunoreactive nerve fibers of the intrahepatic biliary tree were rather sparse in normal livers, intermediate in extrahepatic biliary obstruction and dense in hepatolithiasis. These findings suggest that intrahepatic bile ducts and peribiliary glands are innervated and biliary functions are regulated in part by these nerve fibers. Increased nerve fibers may have altered effects on biliary functions in hepatolithiasis.     

Immunohistochemical demonstration of pancreatic alpha-amylase and trypsin in intrahepatic bile ducts and peribiliary glands

Epithelia of intrahepatic bile ducts and peribiliary glands were immunohistochemically examined for pancreatic alpha-amylase and trypsin in 54 normal autopsied livers. alpha-Amylase was evaluated with a polyclonal antibody, and trypsin was assayed with both polyclonal and monoclonal antibodies. alpha-Amylase was observed in large ducts, septal ducts and peribiliary glands in most livers and was seen in interlobular ducts in seven (13%) livers. Trypsin immunoreactivity with the polyclonal antibody was observed in peribiliary glands in 21 (39%) livers; it was absent in intrahepatic bile ducts in all but one liver. Trypsin immunoreactivity with the monoclonal antibody was present in large ducts, septal ducts and peribiliary glands in about 70% of the livers and was seen in interlobular ducts in two (4%) livers. Bile ductules were always negative for the two antigens. Some epithelia of peribiliary glands positive for both alpha-amylase and trypsin histologically resembled pancreatic acinar cells. alpha-Amylase and trypsin immunoreactivities of intrahepatic biliary epithelia and pancreatic aninar cells were eliminated by absorption of primary antibodies by alpha-amylase or trypsin, suggesting the specificities of the immunoreactivities. These data suggest that epithelia of intrahepatic large ducts, septal ducts and peribiliary glands contain pancreatic alpha-amylase in most livers and that they contain trypsin in about 70% of livers. alpha-Amylase and trypsin may be secreted into intrahepatic bile duct lumens, thereby exerting important effects on the physiology of the intrahepatic biliary tree and hepatic bile.     

The immunobiology of bile and biliary epithelium.

Long thought to be just a simple pipe involved in the delivery of bile from hepatocytes to the gallbladder and intestine, bile ducts are now regarded as highly dynamic structures consisting of cell populations involved in formation, transport and modification of bile by both secretory and absorptive processes. In fact, both bile and biliary epithelium appear to have active immunologic roles in both innate and adaptive immune responses. These roles are becoming increasingly clear as techniques have been developed allowing for the study of bile and biliary epithelial cells (BECs) in mucosal immunity. Bile is actively involved in the transport of immunoglobulin to the intestine, while BECs secrete chemokines and cytokines and serve to localize the immune response by expressing critical cell adhesion molecules. Evidence suggests that BECs may also function as professional antigen-presenting cells (APC) and, in the process, contribute to the modulation of inflammatory reactions. Bile ducts and, in particular, BECs, are the primary site of damage in several immunologically mediated liver diseases. Progress in these important areas has been rapid and forms the basis of this review.     

Intrahepatic bile duct loss in biliary atresia despite portoenterostomy: a consequence of ongoing obstruction?

The histological and immunohistochemical characteristics of the liver in 44 children (28 boys, 16 girls) with extrahepatic biliary atresia at different stages of the clinical course were studied. Thirty-four wedge liver biopsy specimens taken during Kasai operations (25 specimens) and relaparotomy (9 specimens) and 20 hepatectomy explants taken at the time of transplantation were examined. Routine histological stains and monoclonal antibodies against different molecular weight cytokeratins and HLA-DR were used. The histopathological changes and the pattern of cytokeratin expression observed during the course of the disease were suggestive of persistent or recurrent extrahepatic biliary obstruction that occurred despite the Kasai operation and eventually led to cirrhosis and liver failure. Quantitative studies showed a progressive loss of intrahepatic bile ducts over the time course of the disease. This destruction of bile ducts had a geographic anatomical distribution in hepatectomy specimens, and in two livers it occurred predominantly in only one lobe. This geographic distribution of the vanishing bile ducts probably indicates an unpredictable and uneven obliteration of bile ducts in the porta hepatis during portoenterostomy wound healing and scarring.     

Blood group antigens in the intrahepatic biliary tree. I. Distribution in the normal liver

The distribution of six blood group-related antigens, A, B, H, Lea, Leb and sialylated Lea antigens, in the intrahepatic biliary tree was studied. These carbohydrate antigens in fixed normal liver tissues were immunostained with the use of highly specific monoclonal antibodies in an avidin-biotin-peroxidase complex method. Bile ducts expressed both ABH and Lewis blood group antigens. However, only Lewis blood group antigens could be detected in the bile ductules. Canaliculo-ductular junctions could be clearly delineated with the anti-Lewis blood group antibodies, especially with anti-Lea antibody. Small Lewis antigen-positive cells were scattered intralobularly. They were adjacent to parenchymal liver cells, mainly in Rappaport zone 1, and apparently in continuity with the portal biliary tree. Sialylated Lea antigen was found in some septal bile ducts. No blood group antigen could be detected in the bile canaliculi. These results indicate that (1) biliary tract of a given size has its own pattern of blood group antigen expression, and (2) biliary epithelial cells are not identical with regard to the phenotypic expression of their structural carbohydrates. In future, it will be possible to classify biliary epithelial cells by their blood group antigen expression.     

Carcinoembryonic antigen and blood group-related carbohydrate antigens in glycoproteins in human bile in hepatolithiasi

We investigated whether carbohydrate antigens on biliary glycoproteins and carcinoembryonic antigen (CEA) are related to hepatolithiasis. CEA, ABO, and Lewis blood group-related antigens, as well as sialyl-Tn antigen in hepatic bile, were analyzed by Western blotting in samples from 12 patients with hepatolithiasis and 37 with other biliary diseases (choledocholithiasis,13; cholecystolithiasis, 5; acute cholecystitis, 2; cholangiocarcinoma, 5; common bile duct carcinoma, 4; pancreatic carcinoma, 6; and metastatic carcinoma of liver, 2). CEA was positive on mucinous glycoprotein in six patients (50%) with hepatolithiasis and one case (17%) with pancreatic carcinoma. CEA was also positive on a glycoprotein of approximately 200 kd in eight patients (67%) with hepatolithiasis and two (33%) with pancreatic carcinoma. Lewis X was detected on the mucinous glycoprotein in almost all samples, as well as on glycoproteins of approximately 180 kd in all hepatolithiasis samples and approximately half of those from patients with other diseases. Sialyl-Tn antigen was detected on mucinous glycoprotein in four (80%) with cholangiocarcinoma, two (50%) with common bile duct carcinoma, and all pancreatic carcinoma samples. Mucinous glycoprotein and glycoproteins containing CEA and Lewis X antigens are enriched in hepatic bile of hepatolithiasis, and they may be closely related to the formation of intrahepatic calculi. Sialyl-Tn antigen in biliary mucinous glycoprotein will be a good marker of biliary and pancreatic carcinoma, and probably for cholangiocarcinoma complicated with hepatolithiasis. (Hepatology 1996 Feb;23(2):258-63)     

Epithelial cell proliferation activity of the biliary ductal system with congenital biliary malformations

Congenital biliary malformations such as anomalous arrangement of the pancreaticobiliary ductal system (AAPB), congenital cystic dilatation of the common bile duct (CCDB), and congenital biliary strictures at the hepatic hilum (CBSH) are newly designated disease entities and are frequently found in adult patients with biliary malignancy such as gallbladder carcinoma, common bile duct carcinoma, and intrahepatic bile duct carcinoma. In the present study, the relationship of these malformations and biliary malignancy was investigated. We studied 61 gallbladders of patients with AAPB and 56 gallbladders of patients without AAPB; 16 common bile ducts of patients with CCDB (12 with AAPB and 4 without AAPB) and 11 gallbladders of patients without CCDB; and 17 intrahepatic bile ducts of patients with CBSH and 6 intrahepatic bile ducts of patients without CBSH. Tissue sections from the mucosa of the gallbladder, common bile duct, and intrahepatic bile duct were stained for proliferating cell nuclear antigen (PCNA). The PCNA labeling indexes of patients with these malformations were significantly higher than those of patients without these malformations (P < 0.05). Cell proliferation of the epithelia in the biliary ductal system in patients with these congenital biliary malformations was accelerated. Consequently, these congenital malformations appear to be an important risk factor for the occurrence of biliary malignancy.     

Bile duct antibodies crossreacting with blood group antigens in primary sclerosing cholangitis.

Indirect immunoperoxidase histochemistry was used to localise and determine the disease, species, and tissue specificity of bile duct antibodies in primary sclerosing cholangitis. Serum was collected from: 29 patients with primary sclerosing cholangitis, 18 patients with ulcerative colitis alone, 19 patients with extrahepatic biliary obstruction of other causes, and 42 healthy control subjects. Bile duct antibodies reacted with an antigen localised to the small and large intrahepatic bile ducts. When blood group A human liver was used they were detected in 34% of patients with primary sclerosing cholangitis. They were not detected when blood group O human liver was used. Bile duct antibodies that reacted with obstructed and normal rabbit liver were detected in 34% and 17% respectively of patients with primary sclerosing cholangitis but were also present in similar proportions of control subjects. Colon antibodies that reacted with human and rabbit colon were found in 52% and 24% respectively of patients with primary sclerosing cholangitis. Absorption studies using blood group substances A and B abolished the reactivity of bile duct antibodies with human and rabbit liver and that of colon antibodies' with rabbit colon. Colon antibodies that reacted with human colon were not absorbed. Absorption studies using isolated peripheral white blood cells did not affect reactivity of bile duct or colon antibodies. We conclude that bile duct antibodies are disease, species, and tissue non-specific and react with blood group A/B antigens present in human and rabbit bile ducts and rabbit colon. This suggests that they do not play a role in the pathogenesis of primary sclerosing cholangitis.     

Endoscopic retrograde cholangiography in the diagnosis of biliary malformations in infants

The differentiation of infantile biliary malformations from primary parenchymal diseases is difficult. The recent development of a pediatric side-viewing endoscope (PJF Endoscope; Olympus Corporation of America) provided an opportunity to investigate the usefulness of endoscopic retrograde cholangiography (ERC) for precise visualization of the extrahepatic biliary passages in infants with persistent cryptogenic cholestasis. ERC was performed in 12 patients, with visualization of the existing extrahepatic bile ducts in 4. The entire biliary system was visualized in one, excluding extrahepatic biliary atresia and choledochal cyst. The reduced caliber of the intrahepatic bile ducts and histological observations in a percutaneous liver biopsy supported the diagnosis of intrahepatic biliary hypoplasia in this case. An intact hepatic portochole cystostomy was documented in one, although the intrahepatic biliary system was not delineated. Atresia of the hepatic bile ducts proximal to the gallbladder was documented in two. Of the eight patients in whom extrahepatic bile ducts were not visualized by ERC, six had extrahepatic biliary atresia confirmed at exploratory laparotomy. The papilla of Vater could not be located in four of these six infants. The remaining two had neonatal hepatitis. ERC may offer a useful alternative to operative cholangiography in selected infants with persistent cholestasis and acholic stools.     

Heterogeneity of the intrahepatic biliary epithelium

The objectives of this review are to outline the recent findings related to the morphological heterogeneity of the biliary epithelium and the heterogeneous pathophysiological responses of different sized bile ducts to liver gastrointestinal hormones and peptides and liver injury/toxins with changes in apoptotic, proliferative and secretory activities. The knowledge of biliary function is rapidly increasing because of the recognition that biliary epithelial cells (cholangiocytes) are the targets of human cholangiopathies, which are characterized by proliferation/damage of bile ducts within a small range of sizes. The unique anatomy, morphology, innervation and vascularization of the biliary epithelium are consistent with function of cholangiocytes within different regions of the biliary tree. The in vivo models [e.g., bile duct ligation (BDL), partial hepatectomy, feeding of bile acids, carbon tetrachloride (CCl4) orα-naphthylisothiocyanate (ANIT)] and the in vivo experimental tools [e.g., freshly isolated small and large cholangiocytes or intrahepatic bile duct units (IBDU) and primary cultures of small and large murine cholangiocytes] have allowed us to demonstrate the morphological and functional heterogeneity of the intrahepatic biliary epithelium. These models demonstrated the differential secretory activities and the heterogeneous apoptotic and proliferative responses of different sized ducts. Similar to animal models of cholangiocyte proliferation/injury restricted to specific sized ducts, in human liver diseases bile duct damage predominates specific sized bile ducts. Future studies related to the functional heterogeneity of the intrahepatic biliary epithelium may disclose new pathophysiological treatments for patients with cholangiopathies.     

Biliary epithelial cells and primary biliary cirrhosis: the role of liver-infiltrating mononuclear cells

Primary biliary cirrhosis (PBC) is characterized by the highly selective autoimmune injury of small intrahepatic bile ducts, despite widespread distribution of mitochondrial autoantigens. On this basis, it has been suggested that the targeted biliary epithelial cells (BECs) play an active role in the perpetuation of autoimmunity by attracting immune cells via chemokine secretion. To address this issue, we challenged BECs from patients with PBC and controls using multiple Toll-like receptor (TLR) ligands as well as autologous liver-infiltrating mononuclear cells (LMNCs) with subsequent measurement of BEC phenotype and chemokine production and LMNC chemotaxis by quantifying specific chemokines. Our data reflect that BECs from PBC patients and controls express similar levels of TLR subtypes, CD40, and human leukocyte antigen DRalpha (HLA-DRalpha) and produce equivalent amounts of chemokines in our experimental conditions. Interestingly, however, BEC-expressed chemokines elicit enhanced transmigration of PBC LMNCs compared with controls. Furthermore, the addition of autologous LMNCs to PBC BECs led to the production of higher levels of chemokines and enhanced the expression of CD40 and HLA-DRalpha. CONCLUSION: We submit that the proinflammatory activity of BECs in PBC is secondary to the intervention of LMNCs and is not determined per se. These data support the hypothesis that BECs are in fact "innocent victims" of autoimmune injury and that the adaptive immune response is critical in PBC.     

Plasma cells and the chronic nonsuppurative destructive cholangitis of primary biliary cirrhosis

There has been increased interest in the role of B cells in the pathogenesis of primary biliary cirrhosis (PBC). Although the vast majority of patients with this disease have anti-mitochondrial antibodies, there is no correlation of anti-mitochondrial antibody titer and/or presence with disease severity. Furthermore, in murine models of PBC, it has been suggested that depletion of B cells may exacerbate biliary pathology. To address this issue, we focused on a detailed phenotypic characterization of mononuclear cell infiltrates surrounding the intrahepatic bile ducts of patients with PBC, primary sclerosing cholangitis, autoimmune hepatitis, chronic hepatitis C, and graft-versus-host disease, including CD3, CD4, CD8, CD20, CD38, and immunoglobulin classes, as well as double immunohistochemical staining for CD38 and IgM. Interestingly, CD20 B lymphocytes, which are a precursor of plasma cells, were found in scattered locations or occasionally forming follicle-like aggregations but were not noted at the proximal location of chronic nonsuppurative destructive cholangitis. In contrast, there was a unique and distinct coronal arrangement of CD38 cells around the intrahepatic ducts in PBC but not controls; the majority of such cells were considered plasma cells based on their expression of intracellular immunoglobulins, including IgM and IgG, but not IgA. Patients with PBC who manifest this unique coronal arrangement were those with significantly higher titers of anti-mitochondrial antibodies. CONCLUSION: These data collectively suggest a role for plasma cells in the specific destruction of intrahepatic bile ducts in PBC and confirm the increasing interest in plasma cells and autoimmunity.     

Significance of periductal Langerhans cells and biliary epithelial cell‐derived macrophage inflammatory protein‐3α in the pathogenesis of primary biliary cirrhosis

Background/aims: To clarify the primary biliary cirrhosis (PBC)‐specific antigen‐presenting mechanism, we examined the distribution and phenotypic characteristics of infiltrating dendritic cells (DCs) with respect to bile ducts and the mechanism of migration in terms of the periductal cytokine milieu and biliary innate immunity. Methods and results: Immunohistochemistry using liver sections from patients with PBC and controls revealed that blood dendritic cell antigen (BDCA)‐2⁺ plasmacytoid DCs were found mainly in the portal tracts in PBC and the controls, but their distribution was not related to bile ducts. BDCA‐1⁺ and CD19^? myeloid DCs were also found in portal tracts in PBC and the controls and, in particular, Langerin+Langerhans cells (LCs) were dominantly scattered around or within biliary epithelial layers of the damaged bile ducts in PBC. Moreover, experiments with cultured human biliary epithelial cells (BECs) showed that an LC‐attracting chemokine, macrophage inflammatory protein‐3α, was produced by BECs in the response to cytokines [interleukin (IL)‐1β, tumour necrosis factor‐α and IL‐17] and pathogen‐associated molecular patterns. Conclusions: LCs existing around or within biliary epithelial layers are important as periductal antigen‐presenting cells in PBC and the migration of LCs into bile ducts is closely associated with the periductal cytokine milieu and biliary innate immunity in PBC.     

Myeloperoxidase-positive inflammatory cells participate in bile duct damage in primary biliary cirrhosis through nitric oxide-mediated reactions

Previous studies have suggested that increased nitric oxide (NO)-mediated products are found in the livers of subjects with primary biliary cirrhosis (PBC), but the mechanisms involved remain enigmatic. We took advantage of immunohistochemistry and several unique monoclonal antibodies to study inflammatory cells responsible for the generation of NO, the enzymes responsible for NO production, the expression of 3-nitrotyrosine, and the presence of CD68(+) and/or myeloperoxidase (MPO)(+) cells. We examined a total of 113 liver specimens, including 64 with PBC, 19 with primary sclerosing cholangitis (PSC), 6 with non-A, non-B hepatitis, 6 with alcoholic liver disease, 4 with cryptogenic cirrhosis, 4 with biliary atresia, and 10 normal subjects. Twenty-two percent of PBC had elevated expression of 3-nitrotyrosine in their bile duct epithelial cells (BECs) (P =.0316). Furthermore, the BECs in PBC also demonstrated apoptotic changes. MPO-positive inflammatory cells were also noted adjacent to the basement membrane. In contrast, the liver of normal subjects showed few apoptotic changes in the bile ducts, with no evidence of MPO staining in the portal area. Furthermore, sections from livers of subjects with stage I or stage II PBC demonstrated significantly increased inflammatory cell infiltration (P =.0064) and elevated 3-nitrotyrosine expression in BECs (P =.0246) compared with stage III and IV. The presence of 3-nitrotyrosine was closely associated with infiltrating CD68- and/or MPO-positive cells. There was also a stage-associated difference in the presence of bile duct infiltrating cells and 3-nitrotyrosine in PBC with an increase dominant in early stage disease. In conclusion, NO and reactive oxygen species, collectively determined as 3-nitrotyrosine, are associated with bile duct destruction in PBC and are particularly prevalent in early stage disease.     

Immunolocalization of extracellular matrix components and integrins during mouse liver development

Intrahepatic biliary cell differentiation takes place in periportal hepatoblasts under the influence of the subjacent connective tissue, the mechanism of which is still unclear. This study was undertaken to analyze the immunolocalization of extracellular matrix components and their cellular receptors during mouse liver development, with special attention given to biliary differentiation and vascular development. In young fetal mouse liver, primitive structures of sinusoids were developed between hepatic cords associated with hematopoietic cells demonstrated by immunohistochemistry of basal laminar components, the alpha6 integrin subunit, and PECAM-1. Portal veins and hepatic veins showed different staining intensities of alpha2, alpha3, and alpha6 integrin subunits from early stages of development. Anti-beta4 integrin subunit antibodies reacted with portal veins, but not with hepatic veins after perinatal stages. Their different phenotypes may be related to the preferential differentiation of periportal bile ducts. In intrahepatic bile duct development, periportal hepatoblasts adjacent to the connective tissue were immunostained for each basal laminar component on the basal side at almost the same time; alpha3, alpha5, alpha6, and beta4 integrin subunits were immunohistochemically detectable later than the basal laminar components. These staining patterns of intrahepatic bile duct cells clearly differed from those of extrahepatic bile duct cells from the beginning of their development, suggesting that these ducts are of different origins. In conclusion, the vascular structures, including sinusoids, portal veins, and hepatic veins, develop from early stages of liver development, and the extracellular matrix components may play important roles in biliary differentiation and vascular development. Supplementary material for this article can be found on the HEPATOLOGY website (http://interscience.wiley.com/jpages/0270-9139/suppmat/index.html).     

A traumatic neuroma of the bile duct: a case report

Traumatic neuroma of the bile duct is not a true neoplasm, but a reactive proliferation of pericholangial nerve tissue induced by injury. A 60-year-old Japanese man was admitted to investigate obstructive jaundice. He had undergone cholecystectomy and common bile duct exploration 17 years previously. Ultrasonography and computed tomography showed a pneumobilia with dilatation of the intrahepatic biliary ducts. Endoscopic retrograde cholangiography and spiral-computed tomography cholangiography revealed biliary stenosis in the hepatic hilus with dilatation of the intrahepatic biliary ducts. Celiac angiography and arterial portography showed neither tumor stains nor signs of vessel invasion. At surgery, the confluent portion of the intrahepatic biliary ducts in the hepatic hilus was hardly palpable and deformed, but frozen-section microscopic examination confirmed that no malignant cells were present. Anastomosis of the right and left extrahepatic bile duct to the jejunum, reconstructed by Roux-en-Y hepaticojejunostomy, was performed. Histological examination revealed a nodule composed of a haphazard proliferation of nerve fascicles in the fibromuscular layer of the bile duct which were positively stained for S-100 protein. The pathological diagnosis was traumatic neuroma of the bile duct. Thus, the possibility of traumatic neuroma should be considered in the differential diagnosis of patients with late-onset jaundice after biliary tract surgery.     

Differential expression of MHC class II subregion products on bile duct epithelial cells and hepatocytes in patients with primary biliary cirrhosis

To study the expression of MHC Class II subregion gene products on biliary epithelial cells in primary biliary cirrhosis, frozen sections from liver biopsies of 15 patients with primary biliary cirrhosis were studied immunohistochemically using HLA-D subregion specific monoclonal antibodies L243 (HLA-DR), Leu10 (HLA-DQ) and B7/21 (HLA-DP). Patients with early stages of primary biliary cirrhosis showed expression of HLA-DP, HLA-DR and HLA-DQ subregion gene products on bile duct epithelial cells. In advanced stages of disease, no MHC Class II antigens or only HLA-DR and HLA-DP were expressed on bile duct cells. While normal hepatocytes did not express detectable amounts of MHC Class II antigens, hepatocytes from liver biopsies of four patients with primary biliary cirrhosis showed a distinct staining exclusively with monoclonal antibodies specific for HLA-DR. The expression of MHC Class II antigens on parenchymal cells was independent of a lymphocytic infiltration into the tissue. This study demonstrates that bile ductular cells, but not hepatocytes, express a full set of MHC Class II molecules at least during the early stages of primary biliary cirrhosis. We propose, therefore, that the expression of both HLA-DR and HLA-DQ subregion products on bile duct epithelial cells may be a necessary, although not sufficient, condition for the initiation of an autoimmune process leading to the destruction of intrahepatic bile ducts in primary biliary cirrhosis.     

Vascular plexus around intrahepatic bile ducts in normal livers and portal hypertension

Vessels around the intrahepatic large bile ducts (peribiliary vascular plexus) were examined by histologic, immunohistochemical and scanning electron microscopic observations. The vessels within duct walls were mainly capillaries, while those around the duct walls were composed of capillaries and venules. A majority of vessels was positive for factor VIII-related antigen and Ulex europaeus lectin I. Scanning electron microscopy of hepatic arterial and biliary casts revealed that bile ducts were surrounded by the vascular plexus derived from hepatic arterial branches, and serial section observations in addition disclosed the vessels connecting the peribiliary plexus with portal venous branches ('internal roots'). The peribiliary vascular plexus was increased considerably in livers with portal hypertension, especially idiopathic portal hypertension, extrahepatic portal venous obstruction and hepatocellular carcinoma with portal venous tumor thrombi. Internal roots were also frequently found in the livers with portal hypertension. These results suggest that altered intrahepatic hemodynamics in portal hypertensive conditions involves the peribiliary vascular plexus, resulting in an increase of the number and frequent occurrence of 'internal roots', these vessels probably operating as intrahepatic collaterals.